Alcohol administration increases gastrointestinal permeability to bacteria and bacterial endotoxin (lipopolysaccharide (LPS)), and this plays a major role in the initiation of alcohol-induced tissue/organ damage in particular, alcoholic liver disease (ALD). Increased permeability appears to occur principally though the action of the toxic metabolite, acetaldehyde, on interepithelial tight junctions (TJ) that form a major component of the intestinal barrier. Therefore, understanding the underlying mechanisms by which alcohol promotes intestinal epithelial cell (IEC) permeability is important in designing strategies for the prevention or treatment of alcohol-associated medical disorders. In the human body, cells normally grow within a scaffolding of protein and carbohydrate fibers that help create a three dimensional (3-D) structure, thus allowing organs maintain their shape. Difficulties arise when studying cells on Earth as outside of the body, cells tend to grow in flat sheets and are not capable of duplicating the structure they normally hold. Therefore, a 3-D microgravity environment likely represents a more accurate cell culture model of epithelial behavior in vivo. Furthermore, the absence of the fundamental physical force of gravity on epithelial cell barrier function on board the ISS lends a unique opportunity to study the influence of gravity on cellular properties. We hypothesize that the influence of microgravity on the barrier properties of intestinal epithelial cells significantly modifies alcohol-induced effects on epithelial barrier function. We will test this hypothesis in the UH2 phase by (i) quantifying the effects of simulated microgravity on intestinal epithelial cell tight junction proteins and epithelial permeability;(ii) testing the effects of alcohol on barrier properties of IEC under simulated microgravity;(iii) optimizing a 3-dimensional cell culture system to study barrier function on board the ISS. In the UH3 phase we will (iv) quantify the effects of microgravity on IEC permeability induced by alcohol on board the ISS. These studies will provide a definitive answer as to what extent epithelial barrier function is influenced by gravity, and how this impacts upon epithelial responses to ingested toxins. As a result, we will provide new and fundamental knowledge that will likely have significant positive effects on human health, and allow the rational development of new therapeutic strategies for diseases associated with alcohol consumption and deficient intestinal barrier function. PUBLIC HEALTH RELEVANCE: From 2001-2005, there were approximately 79,000 deaths annually attributable to excessive alcohol use, the 3rd leading lifestyle-related cause of death for people in the U.S. each year (Centers for Disease Control). A major contributor to alcohol-induced disease is the ability of alcohol to compromise the normal barrier function of intestinal epithelial cells that line the gut. This project will utilize the unique zero- gravity environment of the International Space Station (ISS) to generate novel fundamental insights into the role of gravity in regulating intestinal barrier properties, and how the absence of gravity modifies the detrimental influence of alcohol on intestinal epithelial cell barrier function.